virioplankton in the kara sea: the impact of viruses on ......borok, nekouzskii raion, yaroslavl...

ISSN 0001�4370, Oceanology, 2015, Vol. 55, No. 4, pp. 561–572. © Pleiades Publishing, Inc., 2015.Original Russian Text © A.I. Kopylov, A.F. Sazhin, E.A. Zabotkina, N.D. Romanova, 2015, published in Okeanologiya, 2015, Vol. 55, No. 4, pp. 620–631.

561

INTRODUCTION

Viruses can be found in all marine ecosystems andare the most abundant component in planktonic com�munities [19, 20, 23]. Viral lysis of planktonic het�erotrophic bacteria can cause more than 60% of bac�terioplankton mortality, considerably affecting thevalues of energy and carbon fluxes, as well as the com�position of bacterial communities in marine ecosys�tems [7, 8, 20, 22]. Sparse studies on ecology of virusesin Arctic waters demonstrate that the abundance ofplanktonic viruses at high latitudes is two orders lowerthat at temperate latitudes [1, 9, 10, 12, 18]. Bacterialmortality caused by viral lysis was determined usingelectron transmission microscopy. It varies from frac�tions of a percent to 40% of bacterioplankton mortal�ity in different regions of the Arctic [2, 12, 17, 18]. Theinformation on the ecology of planktonic viruses in theKara Sea is based only on the results of our studies ofvirioplankton in coastal regions of the sea [3]. All theabove�mentioned facts determine the aim of our study,which is to assess the abundance, spatial distribution,and size structure of virioplankton, the number of bac�terial cells infected by phage viruses, and the virus�induced mortality of bacterioplankton in the pelagialof the Kara Sea.

MATERIALS AND METHODS

The studies were conducted during the 59th cruiseof the research vessel Academician Mstislav Keldysh in

2011. Water samples were collected at stations 5010,5013–5021 along the mouth zone of the Yenisei Riverfrom September 17 to September 22; at stations 5032–5034, 5037, 5039–5042 (the transect through the east�ern slope of the St. Anna Trough (ESSAT)) and at sta�tions 5043–5048 (the transect through the westernslope of the St. Anna Trough (WSSAT) on September28–29 (Figure). Water samples were collected from 2–5 layers using 5�L and 10�L Niskin bottles incorpo�rated into a “Rosette” complex equipped with aCTD�probe (Sea Bird Equipment, United States). Thesampled water layers were determined after the analysisof the temperature, conductivity, and fluorescence.

For the analysis of the total abundance of bacteriawater samples, 25 or 50 mL in volume were fixedimmediately after sampling in a pH�neutral formalde�hyde solution (the final concentration was 1%) andwere stored in polystyrene bottles. The water sampleswere stored at 4°C in the dark. The total abundance ofbacteria was counted using a Leica DM 5000B with×1000 magnification. Bacterial cells were stained withDAPI prior to the analysis [14]. The bacterial wet bio�mass was calculated according to the cell volume bymeans of the ImageScopeColor software. The bacterialbiomass in carbon units was recalculated according tothe cell volume as fg C/cell = 133.754V0.438, wherefg C/cell is the carbon (C) content per one cell, fg; andV is the cell volume, μm3 [3].

Virioplankton in the Kara Sea: the Impact of Viruses on Mortality of Heterotrophic Bacteria

A. I. Kopylova, A. F. Sazhinb, E. A. Zabotkinaa, and N. D. Romanovab

aPapanin Institute for Biology of Inland Waters, Russian Academy of Sciences,Borok, Nekouzskii raion, Yaroslavl oblast, 152742 Russia

e�mail: [email protected]` Institute of Oceanology, Russian Academy of Sciences, Nakhimovskii pr. 36, Moscow, 117997 Russia

Received September 04, 2014

Abstract—Studies were conducted in shallow and deepwater areas of the Kara Sea. The abundance of bacte�ria (NB) and the abundance of viruses (NV) ranged within (19.4–2215.1) × 103 cells/ml and (97.6–5796.8) ×103 particles/ml, respectively. The virus to bacteria ratio varied from 1.4 to 29.1. A positive correlation wasfound between NB and NV (R = 0.87, n = 45, p = 0.05. Using electron transmission microscopy it was detectedthat the frequency of visibly infected cells of bacteria (FVIC) varied from 0.2 to 1.9% of NB. The maximumvalues of FVIC were recorded in the estuary of the Yenisei River. The infected cells of bacteria contained from4 to 127 (an average of 12) phages/cell of mature viruses. Virus�mediated mortality of bacteria was 0.5% andvaried from 1.4 to 16.1% of the total mortality of bacterioplankton. This indicates a minor role of viruses inthe control of overabundance and production of bacterioplankton in the Kara Sea during the surveyed period.

DOI: 10.1134/S0001437015040104

MARINE BIOLOGY

562

OCEANOLOGY Vol. 55 No. 4 2015

KOPYLOV et al.

Bacterial production and bacterioplankton con�sumption by grazers were estimated according to themethod suggested by Sherr et al. [16], using antibioticsin a modification for the natural environment [21].Immediately after water samples were collected, theywere poured into 75�mL sterile transparent polysty�rene bottles; antibiotics were added to some of them.The bottles were placed into large�size mesh containersand were exposed for 8 hours in an aquarium (4.85 ×2.58 × 2.5 m, 31 m3 in volume) with running seawater,located on the upper deck. The period of exposure waschosen based on our experimental data on antibioticactivity dynamics in arctic waters. The container withbottles was placed 1 m below the water surface. Thegrazing of nano� and microphages on bacteria was esti�mated using samples with antibiotics (benzylpenicillin,1 mg/L; vancomycin, 200 mg/L), which stopped bacte�rial growth but had no influence on the grazers [16].Samples without antibiotics were exposed as control.All the sample preparations were performed on thedeck at an air temperature which was close to the tem�perature of the surface water layer. The method of theexperiment is presented in detail in the work by Sazhinet al. [24].

Viral particles were enumerated by epifluores�cence microscopy using SYBR Green I and Anodiscaluminum oxide filters (Wathman) with a pore diam�eter of 0.02 μm [13]. The total amount of countedviral particles was not less than 400 for each filter.The carbon content per one viral particle was takenas 0.055 fg virus–1 [18].

Filters with bacteria and viruses were examinedunder a ×1000 Olympus BX51 (Japan) with a Cell�Fimage�analysis system.

The frequency of visibly infected cells (FVIC), %of the total bacterial number, and the burst size (BS),particles/cell, were determined by transmission elec�tron microscopy. Viruses and bacteria were precipi�tated on 400�mesh pioloform�covered carbon�platednickel grids by centrifugation at 100000 g (35000 rpm)for 2 hours using an OPTIMA L�90k ultracentrifuge(Beckman Coulter, United States). The grids wereexamined under a JEM 1100 (Jeol, Japan) electronmicroscope at magnification ×50000–150000. Oneach grid, not less than 800 bacterial cells were exam�ined. The frequency of infected cells (FIC, % of thetotal number of heterotrophic bacteria) was calculatedaccording to the equation FIC = 7.1FCVI –22.5FVIC2 [6]. The viral�mediated mortality of bacte�ria (VMB, %) was determined according to the for�mula VMB = (FIC + 0.6FIC2)/(1–1.2FIC) [6]. Theabundance of bacterial population was assumed to beconstant, i.e., bacterial production was equal to mor�tality. The rate of virus�induced mortality (VIM),cell/(mL day) or mg C/(m3 day) was calculatedaccording to the equation VIM = VMB × PB, where PB

is the bacterioplankton production. The virioplanktonproduction (PV, particles/(mL day) was calculatedaccording to the equation PV = BS × VIM, where VIM,cell/(mL day). Virus turnover time was determined bythe division of their number by their production. Theamount of easily oxidizable organic matter releasedfrom lysed bacterial cells, mg C/(m3 day) into aquaticenvironment was determined as the differencebetween VIM and РV. These values are, apparently,overestimates, since energy consumption of viruses forthe synthesis of capsid proteins and replication ofnucleic acid was not accounted for. The data on theseissues are not found or are absent in the literature.

The Spearman’s rank coefficient was used to deter�mine the correlation between parameters at a signifi�cance level of 0.05.

RESULTS

The abundance (NB) and biomass (BB) of bacteri�oplankton in the surveyed shallow regions of the KaraSea were considerably higher than in deepwaterregions (Table 1). The values of NB and BB were on

°N

80

79

78

77

76

75

74

73

72

71

7065 70 75 80 85 90 °E

Yamal

Taimyr

Ob

Riv

er

Yen

isei

Riv

erK A R A S E A

Novaya Zemlya

5042

5039

5037

50335045

504850445043

5010

50215019

5018

5013

Diagram of station locations.


VIRIOPLANKTON IN THE KARA SEA 563

average in the water column 2068.6 ± 943.0 ×

103 cell/mL or 41.27 ± 3.23 mg C/m3 in the mouth ofthe Yenisei River, 278.4 ± 89.0 × 103 cell/mL (6.87 ±1.22 mg C/m3) in the estuary, 55.8 ± 11.2 ×

103 cell/mL or 1.97 ± 0.58 mg C/m3 on the easternslope of the St. Anna Trough (ESSAT), and 94.8 ±

39.0 × 103 cell/mL (1.53 ± 0.45 mg C/m3) on the west�ern slope of the St. Anna Trough (WSSAT). A highpositive correlation was determined between watertemperature and NB (R = 0.77, р = 0.05). Experimen�tal estimations of bacterioplankton production dem�onstrated a high reproduction rate of microorganismsat the mouth of the Yenisei River (Table 1). In thelarger estuarine zone the growth of bacteria was notdetected in most of experiments. In the ESSATregion, a relatively high bacterial production wasrecorded only at station 5042, both on the surface layerand at depths of 100 and 461 m; in the near bottomlayer it was 3.85–4.03 mg C/m3 (Table 1). The specificbacterioplankton production (Р/В) was maximal inthe surface layer and near the bottom (2.08 and 2.24,respectively). At other stations in the region PB was low(0.02–1.43 mg C/m3) or almost zero. In the westernpart of the St. Anna Trough, bacterial production wasreliably recorded at all stations and the layers undersurvey except station 5043, where it constituted7.38 mg C/m3 but its values were within the range0.02–0.087 mg C/m3. Р/В varied from 0.13 to 1.31(Table 1).

The abundance of virioplankton (NV) in the waterof the surveyed regions of the sea ranged from 97.6 ×103 particles/mL to 5796.8 × 103 particles/mL, averag�ing 856.6 ± 201.5 × 103 particles/mL (Table 2). Themaximum values of NV in the period under study wererecorded at surface layers, and the minimum valueswere recorded at depths of 150 to 250 m. The abun�dance of viruses at depths lower than 300 m (watertemperature below 0°C) was 1.2–2.1 times higherthan in the upper layers. The ratio NV/NB varied from1.4 to 29.1, averaging 7.1 ± 1.0 (Table 2). A high posi�tive correlation was found between NB and NV (R =0.87, р = 0.05).

The values of NV averaged 4548.5 ± 816.2 × 103 par�ticles/mL near the mouth of the Yenisei River and974.8 ± 422.2 × 103 particles/mL in the estuary of theriver. In the regions of the eastern and western slopesof the St. Anna Trough, NV values were 374.2 ± 37.4 ×106 particles/mL and 479.6 ± 101.6 × 106 parti�cles/mL, respectively. Average values of the NV/NB

ratio were considerably smaller in shallow parts of theestuary of the Yenisei River (2.3 ± 0.5–3.0 ± 0.4) thanin the deepwater parts of the eastern and westernslopes of the St. Anna Trough (10.7 ± 2.0 and 7.9 ± 1.7,respectively).

The number of bacteria with viral particlesattached to their cells (NBV) varied from 1.6 × 103 to330.5 ×103 cells/mL averaging 36.7 ± 11.8 ×

103 cells/mL in the waters of the Kara Sea. The highestvalues were recorded in the region of the Yenisei Rivermouth. The portion of NBV of the total bacterioplank�ton abundance (NBV/NB) varied in a narrower range(6.3–18.8%), averaging 12.7 ± 0.5%. The average val�ues of NBV/NB in shallow parts of the Kara Sea (13.5–15.2%) were close to values in the deepwater regions(11.2–13.0%). From 1 to 30 viral particles weredetected on the surface of one bacterial cell. Thecapsid diameter of attached viruses varied from 18 to184 nm. The number of viruses attached to bacterialcells (NVB) varied from 2.5 × 103 particles/mL to469.6 × 103 particles/mL, averaging 54.2 ± 16.7 ×

103 particles/mL. The NVВ/NV varied within 0.5–33.7% and averaged 6.2 ± 0.9% (Table 2). A high pos�itive correlation was found between NV and NVB: R =0.90, р = 0.05. The average values of NVB and NVВ/NV

were 428.2 ± 38.0 × 103 particles/mL and 9.8 ± 1.0%,respectively, at the mouth of the Yenisei River; 60.5 ±20.6 × 103 particles/mL and 10.5 ± 2.2 in the Yeniseiestuary; 9.3 ± 2.4 × 106 particles/mL and 3.1 ± 0.6% inthe region of the eastern slope of the St. Anna Trough,and 17.0 ± 4.6 × 106 particles/mL and 4.7 ± 1.2% inwaters of the western slope (Table 2).

Viruses without tail appendages predominateamong virioplankton in all surveyed regions of theKara Sea (Table 3). The proportion of such viruses andof viruses with short (to 62 nm) and long (to 560 nm)tail appendages in the total abundance of viruses aver�aged 79.7 ± 1.0%, 13.7 ± 1.0%, and 6.6 ± 0.7%, respec�tively, in all samples.

The diameter of capsids of detected viral particlesvaried from 18 to 389 nm. The average size of capsidsof planktonic viruses varied within 31–89 nm and wason average 72 ± 2 nm (Table 4). In most cases, phagesof 60–100 nm in size were the most numerous groupin the composition of virioplankton (64% of the exam�ined samples). Large phages of 200 to 289 nm in sizewere recorded in 20 water samples (44%), i.e., theirsize was comparable with the size of the smallest bac�terial cells.

As the result, in all surveyed regions of the Kara Seathe contribution of viruses of different size groups 18–40, 40–60, 60–100, 100–150, 150–200 and virusesmore than 200 nm to the total abundance of viri�oplankton was on average 11.9 ± 1.9%, 29.3 ± 1.7%,41.4 ± 1.7%, 13.7 ± 1.1%, 2.7 ± 0.4%, and 1.0 ± 0.2%,respectively.

The frequency of visibly infected cells (FVIC), i.e.,the portion of cells containing mature phage particles inthe total abundance of bacterioplankton varied from 0.2to 1.9% reaching maximum values in the Yenisei mouth

564


KOPYLOV et al.

Table 1. Abundance (NB × 103 cells/mL), biomass (BB mg C/m3), and daily production (PB) of bacterioplankton

No. of station Horizon, m T, °C NB BB

PBP/B

103 cells/mL mg C/m3

Mouth of the Yenisei River

5013 5 9.6 2215.1 47.52 4829 103.60 2.18

15 9.5 2098.2 33.52 3956 63.20 1.89

28 9.5 1892.5 23.62 3674 45.86 1.94

Estuary of the Yenisei River

5018 0 4.7 888.4 14.02 273 4.30 0.31

8 3.6 99.0 3.39 0 0 0

10 2.1 186.9 6.98 0 0 0

20 0.2 121.6 5.06 0 0 0

5019 5 7.0 1045.0 10.93 0 0 0

16 0.9 125.0 6.51 0 0 0

25 –0.5 70.0 5.33 17 1.29 0.24

5021 2 5.5 440.0 16.66 0 0 0

16 0.1 59.0 2.03 0 0 0

23 –0.3 147.0 4.88 0 0 0

5010 5 4.8 142.8 5.42 0 0 0

20 –0.6 136.4 2.36 0 0 0

30 –1.4 157.8 5.73 0 0 0

Eastern slope of the St. Anna Trough

5033 0 4.5 113.5 1.81 0 0 0

9 4.9 140.2 8.97 0 0 0

30 –0.4 19.4 0.74 0 0 0

120 –1.1 28.1 1.69 0 0 0

5037 7 4.1 63.8 3.44 0 0 0

25 1.7 21.1 0.82 37 1.43 1.74

75 0.0 24.3 0.30 28 0.34 1.14

315 –0.3 35.7 0.41 2 0.02 0.06

5039 0 3.4 58.7 1.23 31 0.64 0.52

30 1.3 27.1 0.56 27 0.56 0.99

75 0.5 29.2 0.51 8 0.13 0.26

354 –0.3 27.6 0.98 0 0 0

5042 5 2.9 53.1 1.85 110 3.85 2.08

25 0.9 36.3 0.74 0 0 0

100 1.5 165.3 5.73 116 4.03 0.70

461 –0.4 49.7 1.79 111 4.01 2.24

Western slope of the St. Anna Trough

5043 0 5.3 549.2 6.35 638 7.38 1.16

5044 5 3.3 97.0 2.02 24 0.50 0.25

20 3.5 29.8 0.60 39 0.78 1.31

152 –0.3 44.0 0.95 20 0.43 0.45



Table 1. (Contd.)

No. of station Horizon, m T, °C NB BB

PBP/B

103 cells/mL mg C/m3

5045 0 3.6 80.9 2.66 22 0.72 0.27

20 3.4 58.1 0.83 61 0.87 1.04

100 1.4 27.4 0.53 1 0.02 0.04

527 –0.5 40.5 0.83 17 0.35 0.41

5048 0 4.8 146.9 2.49 3 0.05 0.02

20 3.4 45.9 0.77 10 0.17 0.22

60 3.2 25.0 0.38 6 0.09 0.25

170 –0.3 41.1 0.54 16 0.21 0.40

241 –0.1 46.5 0.89 6 0.11 0.13

and averaging 1.5 ± 0.2%. In the estuary of the YeniseiRiver FVIC averaged 0.6 ± 0.1%, and it was similar inthe regions of the eastern and western slopes of theSt. Anna Trough (0.5 ± 0.1%) (Table 4). The diameterof the capsids of viral particles found in bacterial cellsranged from 15 to 167 nm. A weak positive correlationbetween FVIC and NBV/NB was found in all surveyedwaters (R = 0.26, p = 0.05). The ratio of the number ofbacteria containing distinctly visible viruses to the num�ber of bacteria with viruses attached to the cell surfacewas 7.6–11.5% (on average 10.2 ± 1.3) in the part adja�cent to the Yenisei mouth and 1.4–14.7% (on average4.8 ± 0.5) in other surveyed regions. At the same time, ahigh positive correlation was found between FVIC andNVB (R = 0.65, p = 0.05). Based on estimations of FVICwe calculated that the frequency of infected cells (FIC)in the total bacterioplankton abundance varied within1.4–12.7% (on average 4.1 ± 0.4).

According to our data, heterotrophic bacteria ofdifferent morphologies were infected by viruses to avariable degree. The total number of cells infected byviruses comprised bacilli (54.7%), cocci (29.3%),vibrio (12.0%), and filamentous bacteria (4%).

The phage’s burst size (BS) on average in a samplediffered considerably at different stations and depths,and constituted 12 ± 2 phages/cell. In the surveyedregions the maximum and average BS for all infectedbacteria were 75 and 13 ± 6 phages/cell in the Yeniseimouth, 69 and 14 ± 4 phages/cell in the estuary of theYenisei River, 57 and 9 ± 2 phages/cell in the easternslope of the St. Anna Trough, and 127 and 12 ±

3 phages/cell on the western slope of the St. AnnaTrough.

Virus�induced mortality of bacterioplankton(VMB) during the surveyed period was low and aver�aged 4.5 ± 0.5% in all regions (Table 5). Thus, the aver�age VMB in the part adjacent to the Yenisei mouth

constituted 12.6 ± 1.9%, which considerably exceedsVMB in other surveyed water areas: 4.4 ± 0.7% in theestuary of the Yenisei, 3.7 ± 0.7% in waters of theESSAT, and 4.0 ± 0.5% in the waters of the WSSAT.

The daily virus�induced mortality of bacteria,cells/(mL day) and mg C/(m3 day) (VIM) and the viralproduction (PV) were calculated in the water samplesin which the bacterioplankton production was deter�mined experimentally. The highest VIM values wererecorded at the shallow mouth of the Yenisei River; theVIM values were much lower in the deepwater parts ofthe Kara Sea and decreased with depth (Table 6). As isknown, some amount of organic matter enters theaquatic environment from bacterial cells as the resultof viral lysis. Assuming that the ratio of bacterial pro�duction to their ratio in the waters of the Kara Seaaverages 0.27 [11], we calculated that daily demand ofbacterioplankton for organic matter (СВ) varies within169.8–333.7 mg C/m3 (on average 245.9 ± 47.7) at themouth of the Yenisei River and ranges from 0.3 to27.3 mg C/m3 (on average 5.9 ± 1.1) in other surveyedregions. During the period of our studies in the YeniseiRiver, the amount of organic matter (entering theaquatic environment as the result of viral lysis of bac�teria) which could be repeatedly used by microorgan�isms constituted 2.6–4.3% (on average 3.4 ± 0.5) ofthe daily demand of bacterioplankton for organic mat�ter. In other regions of the Kara Sea the amount ofrepeatedly consumed organic matter constituted 0.4–3.4 (on average 1.2 ± 0.1)% СВ.

Thus, this additional source of nutrients was insig�nificant for bacterioplankton in the Kara Sea duringthe period under survey.

The maximum viral production (PV) and the mini�mum turnover time of virioplankton abundance (ТV)were typical for waters adjacent to the mouth of theYenisei (4158–12026 × 104 h/(mL day) and 0.4–

566


KOPYLOV et al.

Table 2. Abundance of virioplankton (NV), abundance of bacteria with attached viruses (NBV), and abundance of viruses

attached to bacteria (NVB)

No. of station Horizon, m NV, ×103

particles/mL NV/NB

NBV NVB

103 cells/mL % NB 103 h/mL % NV


5013 5 3013.5 1.4 293.5 13.2 352.2 11.7

15 4835.1 2.3 330.5 15.8 462.7 9.6

28 5796.8 3.1 313.1 16.5 469.6 8.1


5018 0 5484.4 6.2 131.9 14.8 184.7 33.7

8 251.6 2.5 11.2 11.4 15.7 6.2

10 485.9 2.6 29.1 15.6 68.0 14.0

20 255.4 2.1 12.6 10.3 34.5 13.5

5019 5 2821.5 2.7 138.5 13.2 256.8 9.1

16 254.2 2.0 16.1 12.8 22.5 8.8

25 374.6 5.4 10.8 15.4 15.1 4.0

5021 2 871.9 2.0 37.6 8.5 48.9 5.6

16 126.4 2.1 11.1 18.8 17.8 14.1

23 409.8 2.8 19.5 13.3 23.4 5.7

5010 5 346.0 2.4 15.7 11.0 20.4 5.9

20 547.3 4.0 19.8 14.5 29.7 5.4

30 441.8 2.8 24.6 15.6 48.6 11.0


5033 0 449.1 4.0 17.8 15.7 26.7 5.9

9 364.5 2.6 12.0 8.6 27.0 7.4

30 565.0 29.1 3.1 16.2 4.3 0.8

120 554.5 20.0 3.8 13.3 4.2 0.8

5037 7 253.7 4.8 6.4 10.0 7.0 4.6

25 407.0 19.3 2.5 12.0 4.2 1.0

75 428.5 17.6 2.7 11.2 3.5 0.8

315 522.2 14.6 5.3 14.8 6.9 5.6

5039 0 173.8 3.0 4.8 8.1 7.7 4.4

30 457.2 16.9 2.1 7.9 2.5 0.5

75 145.8 5.0 2.8 9.5 3.6 2.5

354 289.0 10.5 3.8 13.9 4.6 1.6

5042 5 226.9 4.3 4.2 8.0 5.9 2.6

25 174.1 4.8 3.5 9.7 4.2 2.4

100 371.9 2.3 22.7 13.7 31.8 8.6

461 604.1 12.1 4.2 8.4 5.0 0.8


5043 0 1129.1 2.0 37.1 6.8 48.2 4.3

5044 5 314.7 3.2 15.6 16.1 23.4 7.4

20 163.0 5.5 5.4 18.0 10.8 6.6

152 97.6 2.2 3.1 7.0 4.0 4.1



Table 2. (Contd.)

No. of station Horizon, m NV, ×103

particles/mL NV/NB

NBV NVB

103 cells/mL % NB 103 h/mL % NV

5045 0 935.3 11.6 14.0 17.3 22.4 2.4

20 675.2 11.6 9.0 15.4 14.4 2.1

100 181.5 6.6 3.4 12.2 4.4 2.4

527 385.6 9.5 4.2 10.4 6.3 1.6

5048 0 381.9 2.6 21.4 14.6 53.8 14.1

20 1071.0 23.3 7.7 16.8 10.8 1.0

60 220.9 8.8 1.6 6.3 3.2 1.4

170 570.4 13.9 4.8 11.8 6.2 1.1

241 108.2 2.3 7.4 16.0 13.6 12.6

Table 3. Portion (%) of viruses without tail appendages (1), with short (2) and long (3) tail appendages in total abundanceof virioplankton

Region 1 2 3

Mouth the Yenisei River 74.6–76.7average 75.4 ± 0.7

13.6–14.314.1 ± 0.2

8.9–11.410.5 ± 0.8

Estuary of the Yenisei River 69.8–89.4average 79.6 ± 1.6

6.0–22.113.4 ± 1.6

2.7–12.26.9 ± 0.9


66.7–90.9average 80.7 ± 1.8

0–33.313.9 ± 2.0

0–16.75.4 ± 1.3


67.6–89.4average 80.4 ± 0.8

0–31.213.9 ± 2.5

0–20.05.7 ± 1.5

0.8 days, respectively). In deepwater regions the valuesof PV differed greatly in different layers from 4.2–1294.8 × 104 h/(mL day) in surface layers to 1.8–35 ×104 h/(mL day) at depths more than 30 m. The time ofturnover of the virioplankton abundance ranged from22 hours to 179 days, respectively (Table 6).

DISCUSSION

The analysis of the data demonstrated that thenumber of planktonic viral particles at the mouth ofthe Yenisei River in September 2011 was higher thanin other surveyed regions of the Kara Sea and wassimilar to the abundance of virioplankton in otherarctic coastal waters (Table 7). The number of viri�oplankton in the deepwater parts of the Kara Sea cor�responds to a lower range of values of the abundanceof planktonic viruses in other deepwater regions of

the Arctic (Table 7). According to our and publisheddata there is a positive correlation between the abun�dance of viruses and bacteria [15, 17, 18]. Thevirus/bacteria ratio is smaller in shallow parts of theKara Sea where the average abundance of virioplank�ton is higher than in its deepwater part (1.64 ± 0.53 ×106 and 0.42 ± 0.05 × 106 particles/mL, respectively).In the deepwater part the ratio is greater, i.e.,NV/NB = 2.9 ± 0.3 and 9.4 ± 1.3, respectively.

Viruses with a capsid diameter of 60–100 nm (64%of samples) and 40–60 nm (29% of samples) prevailedamong the virioplankton of the Kara Sea. Accordingto other publications, viral particles with the diameterof capsids in the range from 30–60 nm can dominatein marine waters [5, 20]. It should be noted that viralparticles with the diameter of capsids more than0.2 μm, i.e., with sizes similar to the size of small bac�teria were detected in 20 of 45 analyzed samples. The

568


KOPYLOV et al.

Table 4. Average diameter of the capsid of viral particle (S, nm) and the portion of viral particles with capsids of differentsize in total abundance of virioplankton (%)

No. of station Horizon, m S, nm% of N

V

18–40 40–60 60–100 100–150 150–200 200–389

Mouth the Yenisei River

5013 5 79 ± 5 7.5 26.4 49.1 9.4 1.9 5.7

15 61 ± 3 17.7 46.9 27.8 6.3 0 1.3

28 86 ± 4 7.6 18.1 43.7 22.9 6.7 1.0


5018 0 69 ± 5 7.7 31.4 43.6 12.8 2.6 1.9

8 65 ± 5 20.0 33.3 33.3 13.4 0 0

10 84 ± 7 12.9 25.8 30.6 21.0 6.5 3.2

20 70 ± 4 12.5 25.0 52.1 10.4 0 0

5019 5 79 ± 7 8.5 36.6 36.6 12.7 2.8 2.8

16 69 ± 3 1.2 40.9 48.1 7.4 2.4 0

25 80 ± 9 4.1 13.5 56.7 23.0 2.7 0

5021 2 69 ± 3 9.3 40.8 36.4 9.3 3.4 0.8

16 88 ± 6 6.1 15.2 42.3 30.3 6.1 0

23 86 ± 4 4.3 12.5 37.5 31.9 9.7 4.1

5010 5 89 ± 3 4.3 10.6 53.8 25.9 5.4 0

20 73 ± 4 23.5 19.8 39.5 11.1 4.9 1.2

30 66 ± 3 27.6 29.0 32.9 7.9 2.6 0


5033 0 89 ± 9 0 22.2 51.9 14.8 7.4 3.7

9 69 ± 4 6.0 46.0 38.0 10.0 0 0

30 64 ± 4 6.0 42.0 48.0 4.0 0 0

120 77 ± 4 3.6 28.6 44.6 19.6 3.6 0

5037 7 75 ± 10 10.7 35.7 42.9 7.1 0 3.6

25 54 ± 2 20.0 48.9 31.1 0 0 0

75 80 ± 4 4.3 19.1 53.2 21.3 2.1 0

315 86 ± 10 6.4 28.7 44.6 14.9 4.3 1.1

5039 0 77 ± 4 6.4 12.8 65.9 12.8 2.1 0

30 67 ± 7 11.0 38.9 38.9 5.6 5.6 0

75 76 ± 4 5.6 33.3 36.1 23.6 0 1.4

354 58 ± 8 40.0 33.4 13.3 13.3 0 0

5042 5 74 ± 5 5.6 29.6 49.9 13.0 0 1.9

25 79 ± 7 6.7 25.0 53.3 13.3 1.7 0

100 78 ± 11 13.5 32.4 29.7 20.7 0 3.7

461 75 ± 7 5.3 21.1 52.5 15.8 5.3 0


5043 0 67 ± 3 0 50.0 43.8 6.2 0 0

5044 5 81 ± 8 14.6 18.4 36.6 19.5 7.3 3.6

20 77 ± 8 9.1 31.8 36.4 13.6 9.1 0

152 63 ± 15 42.9 0 42.8 14.3 0 0



Table 4. (Contd.)

No. of station Horizon, m S, nm% of N

V

18–40 40–60 60–100 100–150 150–200 200–389

5045 0 70 ± 4 7.5 37.3 35.8 16.4 3.0 0

20 60 ± 2 11.1 37.0 50.0 1.9 0 0

100 70 ± 7 16.2 41.9 29.7 5.4 5.4 1.4

527 72 ± 7 15.9 32.9 43.2 6.8 0 1.2

5048 0 88 ± 5 0 18.2 52.3 25.0 4.5 0

20 73 ± 4 6.6 19.1 57.1 13.2 3.3 0.7

60 67 ± 5 11.3 38.6 30.2 18.9 0 1.0

170 31 ± 3 72.7 27.3 0 0 0 0

241 69 ± 6 0 42.9 47.6 9.5 0 0

number of large capsids was 2–172 (on average 30 ±10) × 103 particles/mL or 1.61–6.2 (on average 9.2 ±1.0)% of the number of bacterioplankton. Thus, whenenumerating microorganisms by epifluorescencemicroscopy, in some cases large viruses appear likesmall bacterial cells. Based on all available data theaverage diameter of viral capsids was 6.7 times smallerthan the average diameter of a bacterial cell (72 ± 2 and484 ± 16 nm, respectively).

The number of bacteria with viruses attached tothe cell surface was 13.8 ± 0.7% of the bacterioplank�ton abundance in the shallow part of the Kara Seaand 11.5 ± 0.5% in its deepwater parts. Thus, bacte�riophages attacked a rather large number of bacteria.

The number of bacterial cells containing visibleviral particles (1.2–1.9% of the total abundance ofbacteria) and virus�induced mortality of bacteria(9.5–16.1% of the total mortality) at the mouth of theYenisei River were similar to the values which weobtained earlier in the coastal waters of the Kara Sea(1.6 ± 0.2% and 14.1 ± 2.6%, respectively) [2]. In otherregions of the Arctic, the contribution of viruses to thetotal mortality of bacterioplankton was higher andconstituted 9–37% in coastal waters of the Chuckcheeand Bering seas [17] and 6–28% in the Baffin Sea [12].In open parts of the water area in the Kara Sea FVIC(0.2–1.5%, on average 0.5%) and VMB (1.4–10.1%,on average 3.6%) were low.

The FVIC and VMB values obtained in the courseof our studies were close to the values recorded insurface waters of the central Arctic⎯0–1.4% (onaverage 0.5%) and < 1.0–11.0% (on average 4.0%),respectively [18]. The contribution of viral lysis to thetotal mortality of bacterioplankton in deep waters in

the northern part of the Chuckchee Sea varied from2–16% averaging 9% [17]. In the period of our stud�ies the number of mature bacteriophages in bacterialcells in the Kara Sea reached 127 (on average 12 ± 2)phages/cell, which was lower than in the central Arc�tic >200 (on average 35 ± 48) phages/cell [18]. Theanalysis of the size structure of attached and intracel�lular viruses has demonstrated that in the Kara Seaheterotrophic bacteria are infected by bacteriophageviruses with a capsid diameter of 15–184 nm.

The low percentage of virus�infected bacteria iscaused by low abundance of virioplankton and theincrease in water viscosity in cold arctic waters. This,in turn, determines the rate of contacts betweenviruses and hosts which is an order lower in the Arcticthan in temperate waters. For example, such data areobtained for the Baffin Sea where virus�infected bac�teria constitute 6–28% of the total abundance ofmicroorganisms [18].

CONCLUSIONS

A high positive correlation was found between theabundance of virioplankton and bacterioplankton inthe Kara Sea. In the surveyed period, the average size ofcapsids of viral particles was 7 times higher than theaverage size of bacteria. In addition to free viral parti�cles, a large number of viruses were attached to bacte�rial cells. The portion of visibly infected bacterial cellsin the total abundance of bacterioplankton in shallowand deepwater parts and at different stations and layersdiffered greatly. The viral infection of bacteria andvirus�induced mortality of bacterioplankton washigher in the mouth of the Yenisei River than in other

570


KOPYLOV et al.

Table 5. Frequency of visibly infected cells (FVIC, % of NB), frequency of all infected cells of bacteria (FIC, % of NB), bac�terial mortality due to viral lysis (VMB, % of the total mortality), and burst sizes of mature bacteriophages inside infectedcells (BS, phages/cell)

No. of station Horizon, m FVIC FIC VMB BS


5013 5 1.5 10.1 12.3 7 ± 115 1.2 8.2 9.5 32 ± 1728 1.9 12.7 16.1 17 ± 9


5018 0 1.2 8.2 9.5 9 ± 28 0.6 4.2 4.5 45 ± 10

10 0.6 4.2 4.5 8 ± 320 0.2 1.4 1.4 7

5019 5 0.5 3.5 3.7 36 ± 1916 0.7 4.9 5.4 5 ± 0.525 0.6 4.3 4.7 8 ± 1

5021 2 0.2 1.4 1.4 616 1.0 6.9 7.8 35 ± 623 0.5 3.5 3.7 8 ± 4

5010 5 0.5 3.5 3.7 6 ± 220 0.2 1.4 1.4 530 0.7 4.9 5.4 17 ± 8


5033 0 0.4 2.8 3.0 14 ± 109 0.4 2.8 3.0 8 ± 4

30 0.5 3.5 3.7 11 ± 5120 0.5 3.5 3.7 9 ± 5

5037 7 0.2 1.4 1.4 525 1.5 10.1 12.3 35 ± 975 0.3 2.1 2.2 9 ± 0.5

315 0.6 4.2 4.5 10 ± 6

5039 0 0.3 2.1 2.2 6 ± 130 0.4 2.8 3.0 7 ± 0.375 0.4 2.8 3.0 9 ± 1

354 0.9 6.2 7.0 7 ± 1

5042 5 0.8 5.5 6.1 8 ± 225 0.2 1.4 1.4 7

100 0.4 2.8 3.0 10 ± 3461 0.2 1/4 1/4 5


5043 0 1.0 6.9 7.8 26 ± 9

5044 5 0.4 2.8 3.0 820 0.5 3.5 3.7 10 ± 2

152 0.4 2.8 3.0 9

5045 0 0.4 2.8 3.0 18 ± 420 0.7 4.9 5.4 48 ± 39

100 0.2 1.4 1.4 7527 0.2 1.4 1.4 9

5048 0 0.4 2.8 3.0 14 ± 320 0.5 3.5 3.7 25 ± 760 0.8 5.5 6.1 8 ± 1

170 0.8 5.5 6.1 17 ± 6241 0.5 3.5 3.7 8 ± 2



parts of the Kara Sea. In the deepwater part of the KaraSea relatively high values of these parameters werecharacteristic both of surface and deep waters. Gener�ally, bacteriophages played a minor role in the controlover abundance and production of heterotrophic bac�terioplankton in the period under survey.

ACKNOWLEDGMENTS

The work was supported by the Russian Foundationfor Basic Research, projects nos. 14�04�00130a and

14�05�10055_K and the Russian Scientific Founda�tion, project no. 14�50�00095, processing of materials.

REFERENCES

1. M. P. Venger, T. I. Shirokolobova, P. R. Makarevich,and V. V. Vodop’yanova, “Viruses in the pelagic zone ofthe Barents Sea,” Dokl. Biol. Sci. 446 (1), 306–309(2012).

2. A. I. Kopylov, D. B. Kosolapov, E. A. Zabotkina,P. V. Boyarskii, V. N. Shumilkin, and N. A. Kuznetsov,“Planktonic viruses, heterotrophic bacteria, and

Table 6. The number of lysed bacteria (VIM), production of viruses (Pv × 104 h/(mL day)) and the turnover time of viruses

(T, day)

Depths, m; number of measurements, ( )

VIMPv

T103 cells/(mL day) µm C/(m3 day)


0–28 (3) 375.8–594.0average 520.4 ± 72.3

6004–127428710 ± 2055

4158–120268756 ± 2369

0.4–0.8 0.6 ± 0.1

Estuary of the Yenisei River and the St. Anna Trough

0–25 (11) 0.4–49.8average 8.7 ± 4.7

6–576136 ± 54

4.2–1294.8275.9 ± 83.2

0.9–107.132.8 ± 13.0

30–100 (5) 0.3–3.5average 1.1 ± 0.6

4–12131 ± 22

2.7–35.010.2 ± 6.2

10.6–93.3 61.3 ± 14.2

152–241 (3) 0.2–1.0average 0.6 ± 0.2

4–1310 ± 3.0

1.8–17.08.1 ± 4.6

18.1–60.1 37.2 ± 12.3

461–527 (2) 0.2–1.6average 0.9

5–5630

2.2–8.05.1

75.5–178.5127.0

Table 7. Abundance of bacterioplankton (NB × 105 cells/mL) and abundance of virioplankton (NV × 106 particles/mL) in

the surface waters of the Arctic

Area of studies Month NB NV

NV/NB, % Source

Coastal waters

Beaufort Sea 05–08, 10 0.9–33.7 0.2–10.9 0.85–0.0 Clasen et al. [7]

Kara Sea 08–09 10.5–30.6 3.3–25.1 2.3–9.9 Kopylov et al. [2]

Kara Sea, mouth of the Yenisei River 09 18.92–22.15 3.01–5.80 1.4–3.1 Present study

Open waters

Central Arctic 09–10 0.03–0.47 0.68–11.0 5–70 Steward et al. [18]

Barents sea 06–07 4.1–60.0 0.7–9.0 3 (average) Howard�Jones et al. [9]

Barents sea 09 0.1–4.4 1.7–64.1 2.4–43.5 Venger et al. [1]

Chuckchee Sea 08 0.8–8.7 0.1–1.7 – Hodges et al. [8]

Chuckchee Sea, northern part 08–09 2.1 2.5 – Steward et al. [17]

Kara Sea, eastern part 09 0.6–10.4 0.1–5.4 2.0–6.2 Present study

Kara Sea, St. Anna Trough 09 0.25–.5 0.1–1.1 2.0–29.1 Present study

572


KOPYLOV et al.

nanoflagellates in fresh and coastal marine waters ofthe Kara Sea Basin (the Arctic),” Inland Water Biol. 5(3), 241–249 (2012).

3. N. D. Romanova and A. F. Sazhin, “Relationshipsbetween the cell volume and the carbon content of bac�teria,” Oceanology (Engl. Transl.) 50 (4), 522–530(2010).

4. A. F. Sazhin, N. D. Romanova, and S. A. Mosharov,“Bacterial and primary production in the pelagic zoneof the Kara Sea,” Oceanology (Engl. Transl.) 50 (5),759–765 (2010).

5. M. C. Alonso, G. F. Jimenez, T. Rodriguez, andJ. J. Borrego, “Distribution of virus�like particles in anoligotrophic marine environment (Alboran Sea, WesternMediterranean),” Microb. Ecol. 42, 407–415 (2001).

6. B. Binder, “Reconsidering the relationship betweenvirally induced bacterial mortality and frequency ofinfected cells,” Aquat. Microbial. Ecol. 18, 207–215(1999).

7. J. L. Clasen, S. M. Brigden, J. P. Payet, and C. A. Sut�tle, “Evidence that viral abundance across oceans andlakes is driven by different biological factors,” Freshwa�ter Biol. 53, 1090–1100 (2008).

8. L. R. Hodges, N. Bano, J. T. Hollibaugh, and P. Yager,“Illustrating the importance of particulate organic mat�ter to pelagic microbial abundance and communitystructure – an Arctic case study,” Aquat. Microb. Ecol.40, 217–227 (2005).

9. M. H. Howard�Jones, V. D. Ballard, A. E. Allen, et al.,“Distribution of bacterial biomass and activity in themarginal ice zone of the central Barents Sea duringsummer,” J. Mar. Syst. 38, 77–91 (2002).

10. R. Maranger, D. F. Bird, and S. K. Juniper, “Viral andbacterial dynamics in arctic sea ice during the springalgal bloom near Resolute, NWT, Canada,” Mar. Ecol.:Progr. Ser. 111, 121–127 (1884).

11. B. Meon and R. M. W. Amon, “Heterotrophic bacterialactivity and fluxes of dissolved free amino acids and glu�cose in the Arctic rivers Ob, Yenisei, and the adjacentKara Sea,” Aquat. Microb. Ecol. 37, 121–135 (2004).

12. M. Middelbore, T. G. Nielsen, and P. K. Biorsen,“Viral and bacterial production in the North Water insitu measurements batch�culture experiments and

characterization of a viral�host system,” Deep�SeaRes. 49, 5063–5079 (2002).

13. R. T. Noble and J. A. Fuhrman, “Use of SYBR Greenfor rapid epifluorescence count of marine viruses andbacteria,” Aquat. Microb. Ecol. 14, 113–118 (1998).

14. K. G. Porter and Y. S. Feig, “The use DAPI for identi�fying and counting of aquatic microflora,” Limnol.Oceanogr. 25 (5), 943–948 (1980).

15. C. Säwström, J. Laybourn�Parry, W. Graneli, andA. M. Anesio, “Heterotrophic bacterial and viraldynamics in Arctic freshwaters: results from a fieldstudy and nutrient�temperature manipulation experi�ments,” Polar Biol. 30, 1407–1415 (2007).

16. B. F. Sherr, E. B. Sherr, T. L. Andrew, R. D. Fallon,and S. Y. Newell, “Trophic interactions between het�erotrophic Protozoa and bacterioplankton in estua�rine water analyzed with selective metabolic inhibi�tors,” Mar. Ecol.: Progr. Ser. 32, 169–179 (1986).

17. G. F. Steward, D. C. Smith, and F. Azam, “Abundanceand production of bacteria and viruses in the Beringand Chukchi seas,” Mar. Ecol.: Progr. Ser. 131, 287–300 (1996).

18. G. F. Steward, L. B. Fandino, J. T. Hollibaugh,T. E. Whitledge, and F. Azam, “Microbial biomass andviral infections of heterotrophic prokaryotes in the sub�surface layer of the central Arctic Ocean,” Deep SeaRes., Part I 54, 1744–1757 (2007).

19. C. A. Suttle, “Viruses in the sea,” Nature 437, 356–361(2005).

20. M. G. Weinbauer, “Ecology of prokaryotic viruses,”FEMS Microbiol. Rev. 28 (2), 127–181 (2004).

21. T. Weisse, “The microbial loop in the Red Sea: dynam�ics of pelagic bacteria and heterotrophic nanoflagel�lates,” Mar. Ecol.: Progr. Ser. 55, 241–250 (1989).

22. L. E. Wells and J. W. Deming, “Significance of bacte�rivory and viral lysis in bottom waters of Franklin Bay,Canadian Arctic, during winter,” Aquat. Microb. Ecol.43, 209–221 (2006).

23. K. E. Wommack and R. R. Colvell, “Viruses in aquaticecosystems,” Microbiol. Mol. Biol. Rev. 64, 69–114(2000).

Translated by N. Ruban

virioplankton in the kara sea: the impact of viruses on ......borok, nekouzskii raion, yaroslavl...

Documents